Hematopoietic progenitor cell mobilization using low‐dose cyclophosphamide and granulocyte colony‐stimulating factor for multiple myeloma

High‐dose chemotherapy (HDT) supported by autologous stem cell transplantation (ASCT) has long been one of the standards of care for younger patients with multiple myeloma (MM). Cyclophosphamide (CY) plus granulocyte colony‐stimulating factor (G‐CSF) has been the conventional preparation for hematopoietic progenitor cell (HPC) mobilization, although the optimal dosage of CY in this setting has not yet been clearly defined. This study investigated the efficacy and safety of low‐dose (LD‐)CY (1.5 g/m²) plus G‐CSF for conditioning for HPC apheresis harvest (HPC‐A) in 18 MM patients, and compared it with a regimen consisting of intermediate‐dose (ID)‐CY (4 g/m²) plus G‐CSF for 13 MM patients. Eleven patients in the former and six in the latter were treated with bortezomib (BTZ) during the induction therapy. Both regimens were comparably effective in terms of CD34⁺ cell yields, while adverse events, such as leukopenia, thrombocytopenia, and febrile neutropenia, occurred significantly less frequently in the LD‐CY cohort. All patients in LD‐CY cohort started and completed their apheresis on day 7 or 8, whereas for the ID‐CY cohort the day of first apheresis varied widely from day 8 to 15. These findings indicate that the LD‐CY regimen is as effective as ID‐CY for HPC mobilization, while the former is clearly more practicable and convenient than the ID‐CY regimen for patients with MM. J. Clin. Apheresis 28:368–373, 2013. © 2013 Wiley Periodicals, Inc.     

Alteration of adipokines during peripheral blood stem cell mobilization induced by granulocyte colony‐stimulating factor

Adipokines, soluble mediators produced by adipocytes, have been shown to play a role in various physiological and pathological conditions. We investigated the involvement of adipokines in granulocyte colony‐stimulating factor (G‐CSF)‐induced mobilization of hematopoietic stem cells in 21 healthy donors. We found that serum visfatin and resistin levels, but not leptin and adiponectin levels, were significantly elevated by G‐CSF treatment. G‐CSF treatment activated signaling proteins like extracellular signal‐regulated kinase and stimulated secretion of visfatin from 3T3‐L1 adipocytes. These findings suggest that some adipokines may play a role in G‐CSF‐induced mobilization of stem cells from the bone marrow into systemic circulation. J. Clin. Apheresis 2009. © 2009 Wiley‐Liss, Inc.     

Predictors of unsuccessful mobilization with granulocyte colony‐stimulating factor alone in patients undergoing autologous hematopoietic stem cell transplantation

Mobilization of hematopoietic stem cells is achieved with hematopoietic growth factors with or without chemotherapy or other agents. Although studies comparing granulocyte colony‐stimulating factor (G‐CSF) alone to combined regimens demonstrate an increase in stem cell yield in the latter, mobilization with G‐CSF alone is still effective and has been widely practiced. We conducted a retrospective cohort study of consecutive patients at our institution who underwent at least one mobilization attempt with G‐CSF between January 2000 and December 2008 to identify the proportion of patients failing one or more mobilization attempts and the potential predictors of mobilization failure with this regime. Out of 293 patients, 251 (86.6%) were successfully mobilized and 244 (83.6%) underwent hematopoietic stem cell transplantation. Median yield was 3.55 × 10⁶ CD34⁺ cells/kg. On univariate analysis, mobilization success was influenced by degree of previous treatment and underlying diagnosis (P < 0.001 each) but not by age (P = 0.114), sex (P = 0.860), or radiotherapy (P = 0.454). A diagnosis of non‐Hodgkin's lymphoma (NHL) and number of previous chemotherapy regimens were predictors of failure on multivariate analysis. CD34⁺ yield was influenced by diagnosis and previous chemotherapy (P < 0.001 each). Mobilization with G‐CSF alone yields adequate collections for most patients; however, heavily pretreated NHL patients with one failed attempt had high rates of remobilization failure and should be considered for alternative regimens. J. Clin. Apheresis 28:285–292, 2013. © 2013 Wiley Periodicals, Inc.     

Pulmonary granulocyte kinetics in relation to endothelial and granulocyte activation

The aim of the study was to measure the peripheral blood levels of soluble E-selectin in patients with systemic inflammation and compare them with in vivo granulocyte activation, pulmonary intravascular granulocyte pooling, pulmonary extravascular granulocyte migration and 99mTc-diethylenetriaminepenta-acetic acid (DTPA) aerosol clearance, an index of lung injury. The level of soluble E-selectin was measured by capture ELISA. Granulocytes were labelled with 111In and 99mTc for quantification of pulmonary granulocyte kinetics. The pulmonary vascular granulocyte pool (PGP) was expressed as a fraction of the total blood granulocyte pool. Pulmonary granulocyte migration was quantified on 24-h images using the 111In signal. Granulocyte activation was quantified as the percentage of circulating cells showing shape change ('primed'). Lung injury was assessed from the clearance rate of inhaled 99mTc-DTPA aerosol. Eighteen patients with systemic inflammation were studied: five with inflammatory bowel disease, eight with systemic vasculitis, four with graft versus host disease and one with a recent renal transplant. The peripheral blood levels of soluble E-selectin were significantly elevated in patients with systemic inflammation. The level of soluble E-selectin showed a significant association with granulocyte migration (Spearman rank correlation coefficient, Rs=0.53; P<0.05) but not with PGP or with the percentage of cells showing shape change (P>0.05 for both). Granulocyte migration was bimodal: patients were therefore subdivided into 'migrators' and 'non-migrators'. Soluble E-selectin level, 99mTc-DTPA clearance and PGP, but not the percentage of cells showing shape change, were significantly higher in migrators than in non-migrators. We conclude that pulmonary intravascular granulocyte pooling is increased in the presence of increased numbers of circulating primed granulocytes but increased pooling does not by itself promote granulocyte migration into the lung interstitium. Insofar as an elevated level of E-selectin in peripheral blood reflects vascular endothelial activation, the data are consistent with the notion that pulmonary endothelial activation is required, in addition to granulocyte activation and an expanded PGP, for granulocyte migration into lung parenchyma and, therefore, for lung injury to occur.     

Effects of granulocyte–colony-stimulating factor on potential normal granulocyte donors

BACKGROUND: The use of granulocyte-colony-stimulating factor (G-CSF) to increase the granulocyte count and the yield from leukapheresis in normal donors is leading to renewed interest in granulocyte transfusion. Therefore, it is important to understand the side effects of G-CSF. STUDY DESIGN AND METHODS: We studied the effect of G-CSF on peripheral blood counts and recorded the side effects experienced 24 hours after an injection of G-CSF in normal subjects donating peripheral blood progenitor cells for research. RESULTS: Following administration of G-CSF to 261 donors, the neutrophil count increased to 20.6 to 24.5 x 10(9) per microL depending on the dose of G-CSF. This represented a 6.2 to 7.4-fold increase over the neutrophil count before G-CSF administration. Of all donors, 69 percent experienced one or more side effects. The most common effects were: muscle and bone pain, headache, fatigue, and nausea. There was a relationship between the dose of G-CSF and the likelihood of experiencing a side effect. Most side effects were mild, but about 75 percent of donors took analgesics because of them. CONCLUSIONS: In a granulocyte donation program involving G-CSF stimulation, about two-thirds of donors would experience one or more side effects, but these would usually be mild and well tolerated.     

A study of granulocyte cytotoxins and detection of granulocyte allospecific antigens

Granulocytes were isolated b overlaying leukocyte-rich plasma discontinuous density gradients. The cell suspensions were 95 percent granulocytes. Sera of 383 polytransfused patients and 521 multiparous women were examined for lymphocytotoxins, granulocytotoxins, and granulocytoagglutinins. On the basis of these tests, sera were identified which contained only granulocytotoxins. These were further identified with more precision as to their titers and antibody absorption; reactivity with granulocytes, B, and T lymphocytes; and K 562 cell line. On the basis of calculated correlation coefficients, nine sera were selected which determined three granulocyte specificities having genotype frequencies consistent with the Hardy-Weinberg equilibrium.     

Eighteen years experience of granulocyte donations—acceptable donor safety?

Background: Granulocyte transfusions are given to patients with life‐threatening infections, refractory to treatment. The donors are stimulated with corticosteroids ± granulocyte colony stimulating factor (G‐CSF). However, data regarding the donors' safety is sparse. The objective was therefore to evaluate short‐ and long‐term adverse events (AE) in G‐CSF stimulated donors. Study design and methods: All consecutive granulocyte donors from 1994 to 2012 were identified through our registry. From the donation records, the number of aphereses, stimulation therapy, AE, blood values post donation, and recent status were evaluated. Results: One hundred fifty‐four volunteer donors were mobilized for 359 collections. Age at first granulocyte donation was 43 years (median; range 19–64 years). Follow‐up was 60 months (median; range 0–229 months). The dose of G‐CSF per collection was 3.8 ug/kg body weight (median; range 1.6–6.0 ug/kg). Sedimentation agent was HES. Short‐term AE were mild. Blood values 4 weeks post donation with minor reductions/elevations mostly resolved in later donations. Fourteen donors were excluded from the registry due to hypertension (4), diabetes (2), atrial flutter (1), breast carcinoma (1), urethral carcinoma in situ (1), MGUS (1), thrombosis (1), anaphylaxis (1), primary biliary cirrhosis (1), and unknown (1). Three donors are deceased due to diabetes, acute myocardial infarction, and unknown cause. All excluded/deceased donors except one were excluded/died at least 6 months after first granulocyte donation. Conclusion: No serious short‐term AE were observed. Due to the variability of diagnoses among excluded/deceased donors, we propose that it is less likely that granulocyte donations have a causative impact on these donors' exclusion or death. J. Clin. Apheresis 30:265–272, 2015. © 2014 Wiley Periodicals, Inc.     

Endothelial cells suppress granulocyte clusters and stimulate large granulocyte colonies in culture

Endothelial cells have been shown to produce granulopoietic colony stimulating activity (CSA) under the regulatory control of a humoral factor, MRA produced by blood monocytes. An endothelial cell-derived granulopoietic inhibitory factor has also been described. To further define these apparently paradoxical observations, human bone marrow mononuclear cells were co-cultured with umbilical cord derived endothelial cells in a plasma clot system in vitro. To enhance the sensitivity of the assay for growth effects attributable to the endothelial cells (or their products) alone, an exogenous source of CSA (e.g. a peripheral blood leukocyte feeder layer) was not used. On day ten of culture, less than or equal to 1% endothelial cells markedly stimulated the growth of early granulocyte progenitors (large diaminofluorine positive (DAF+) GM-CFUc) (p less than .01) and a linear dose response relationship was confirmed (p less than .001). Late granulocyte progenitors (DAF+ clusters) were coincidently suppressed by less than or equal to 2% endothelial cells (p less than .01). No effect of endothelial cells on intermediate progenitors (small GM-CFUc) was demonstrated at any concentration. Similar effects were observed with the addition of 5% to 30% endothelial conditioned medium (ECM) (p less than .01). When cohort cultures were evaluated serially, suppression of clusters was observed by day four and stimulation of large GM-CFUcs by day six. These varied effects on different stages of granulocyte differentiation suggest that endothelial cell derived CSA(S) may be of biological relevance in the regulation of granulopoiesis.     

In vitro granulocyte aggregation

Granulocyte aggregation in EDTA-blood from a patient with rheumatoid arthritis is described. The phenomenon was observed three times within a 10-month period. The blood leukocyte count in a Coulter Counter decreased approximately 30% within 6 h, without resulting in overt leukopenia. Simultaneously, increasing numbers of granulocyte aggregates turned up in smears. There were no indications of thrombocytes being involved in the phenomenon.     

Studies on granulocyte preservation. III. Effect of agitation on granulocyte concentrates

The effect of agitation on granulocyte storage was examined. Granulocyte concentrates were obtained as buffy coats from fresh blood by centrifugation and stored for up to 48 hours at 22 degrees C with or without horizontal agitation (80 rpm). The cell counts, mean cell volumes, morphologic changes, phagocytosis, and bactericidal activity of the stored granulocytes did not differ significantly. However, chemotaxis was maintained better in granulocytes that were agitated. At 48 hours, the ability of unagitated cells to adhere to both immunoglobulin-coated and uncoated glass surfaces increased, and clumps were observed on the surfaces. These results seem to be related to the decrease in chemotaxis. The pH of unagitated sedimented cells was more acid. To avoid the decrease of this local pH, stored granulocytes need gentle agitation.